Bacillus subtilis, known also as the hay bacillus or grass bacillus, is a Gram-positive, catalase-positive bacterium, found in soil and the gastrointestinal tract of ruminants, humans, and marine sponges.
Bacillus subtilis is motile and amylase-positive. It forms biofilms through the formation of extracellular polymeric matrix containing sugars and proteins.
As a member of the genus Bacillus, B. subtilis is rod-shaped and can form a tough, protective endospore, allowing it to tolerate extreme environmental conditions. It historically has been classified as an obligate aerobe, though evidence exists that it is a facultative anaerobe. B. subtilis is considered the best-studied Gram-positive bacterium and a model organism to study bacterial chromosome replication and cell differentiation. It is one of the bacterial champions in secreted enzyme production and used on an industrial scale by biotechnology companies.
Description Bacillus subtilis is a Gram-positive bacterium, rod-shaped and catalase-positive. It was originally named Vibrio subtilis by Christian Gottfried Ehrenberg, and renamed Bacillus subtilis by Ferdinand Cohn in 1872 (subtilis being the Latin for "fine, thin, slender"). B. subtilis cells are typically rod-shaped, and are about 4–10 micrometers (μm) long and 0.25–1.0 μm in diameter, with a cell volume of about 4.6 fL at stationary phase.
As with other members of the genus Bacillus, it can form an endospore to survive extreme environmental conditions of temperature and desiccation. B. subtilis is a facultative anaerobe and had been considered as an obligate aerobe until 1998. B. subtilis is heavily flagellated, which gives it the ability to move quickly in liquids.
B. subtilis has proven highly amenable to genetic manipulation and has become widely adopted as a model organism for laboratory studies, especially of sporulation, which is a simplified example of cellular differentiation. In terms of popularity as a laboratory model organism, it is often considered as the Gram-positive equivalent of Escherichia coli, an extensively studied Gram-negative bacterium. The Indian State of Kerala declared the microbe as its own State microbe.
Characteristics Colony, morphological, physiological, and biochemical characteristics of B. subtilis are shown in the table below.
Test type Test CharacteristicsColony charactersSizeMediumTypeRoundColorWhitishShapeConvexMorphological charactersShapeRoundPhysiological charactersMotility +Growth at 6.5% NaCl+Biochemical charactersGram staining+Oxidase-Catalase+Oxidative-FermentativeFermentativeMotility-+Methyl Red-Voges-Proskauer++Indole-H2S Production+Urease-Nitrate reductase+β-Galactosidase+Hydrolysis ofGelatin+Aesculin+Casein+Tween 40+Tween 60+Tween 80+Acid production fromGlycerol+Galactose+D-Glucose+D-Fructose+D-Mannose+Mannitol+N-Acetylglucosamine+Amygdalin+Maltose+D-Melibiose+D-Trehalose+Glycogen+D-Turanose+ Note: + = Positive, – = Negative
Habitat This species is commonly found in the upper layers of the soil and is thought to be a normal gut commensal in humans. A 2009 study compared the density of spores found in soil (about 106 spores per gram) to that found in human feces (about 104 spores per gram). The number of spores found in the human gut was too high to be attributed solely to consumption through food contamination. In some bee habitats, B. subtilis appears in the gut flora of honey bees. B. subtilis can also be found in marine environments.
Some evidence indicates that B. subtilis is saprophytic in nature. The bacterium exhibits vegetative growth in soil rich in organic matter, and that spores were formed when nutrients were depleted. Additionally, B. subtilis has been shown to form biofilms on plant roots, which might explain why it is commonly found in gut microbiomes. Perhaps animals eating plants with B. subtilis biofilms can foster growth of the bacterium in their gastrointestinal tracts. The entire lifecycle of B. subtilis can be completed in the gastrointestinal tract, which lends credence to the idea that the bacterium enters the gut via plant consumption and stays present as a result of its ability to grow in the gut.
Reproduction
Bacillus subtilis can divide symmetrically to make two daughter cells (binary fission), or asymmetrically, producing a single endospore that can remain viable for decades and is resistant to unfavourable environmental conditions such as drought, salinity, extreme pH, radiation, and solvents. The endospore is formed at times of nutritional stress and through the use of hydrolysis, allowing the organism to persist in the environment until conditions become favourable. Prior to the process of sporulation the cells might become motile by producing flagella, take up DNA from the environment, or produce antibiotics. These responses are viewed as attempts to seek out nutrients by seeking a more favourable environment, enabling the cell to make use of new beneficial genetic material or simply by killing off competition.
Under stressful conditions, such as nutrient deprivation, B. subtilis undergoes the process of sporulation. This process has been very well studied and has served as a model organism for studying sporulation.
Sporulation
Once B. subtilis commits to sporulation, the sigma factor sigma F is secreted. This factor promotes sporulation. A sporulation septum is formed and a chromosome is slowly moved into the forespore. When a third of one chromosome copy is in the forespore and the remaining two thirds is in the mother cell, the chromosome fragment in the forespore contains the locus for sigma F, which begins to be expressed in the forespore. In order to prevent sigma F expression in the mother cell, an anti-sigma factor, which is encoded by spoIIAB, is expressed. Any residual anti-sigma factor in the forespore (which would otherwise interfere with sporulation) is inhibited by an anti-anti-sigma factor, which is encoded by spoIIAA. SpoIIAA is located near the locus for the sigma factor, so it is consistently expressed in the forespore. Since the spoIIAB locus is not located near the sigma F and spoIIAA loci, it is expressed only in the mother cell and therefore repress sporulation in that cell, allowing sporulation to continue in the forespore. Residual spoIIAA in the mother cell represses spoIIAB, but spoIIAB is constantly replaced so it continues to inhibit sporulation. When the full chromosome localizes to the forespore, spoIIAB can repress sigma F. Therefore, the genetic asymmetry of the B. subtilis chromosome and expression of sigma F, spoIIAB and spoIIAA dictate spore formation in B. subtilis.
Chromosomal replication
Bacillus subtilis is a model organism used to study bacterial chromosome replication. Replication of the single circular chromosome initiates at a single locus, the origin (oriC). Replication proceeds bidirectionally and two replication forks progress in clockwise and counterclockwise directions along the chromosome. Chromosome replication is completed when the forks reach the terminus region, which is positioned opposite to the origin on the chromosome map. The terminus region contains several short DNA sequences (Ter sites) that promote replication arrest. Specific proteins mediate all the steps in DNA replication. Comparison between the proteins involved in chromosomal DNA replication in B. subtilis and in Escherichia coli reveals similarities and differences. Although the basic components promoting initiation, elongation, and termination of replication are well-conserved, some important differences can be found (such as one bacterium missing proteins essential in the other). These differences underline the diversity in the mechanisms and strategies that various bacterial species have adopted to carry out the duplication of their genomes.
Genome
Bacillus subtilis has about 4,100 genes. Of these, only 192 were shown to be indispensable; another 79 were predicted to be essential, as well. A vast majority of essential genes were categorized in relatively few domains of cell metabolism, with about half involved in information processing, one-fifth involved in the synthesis of cell envelope and the determination of cell shape and division, and one-tenth related to cell energetics.
The complete genome sequence of B. subtilis sub-strain QB928 has 4,146,839 DNA base pairs and 4,292 genes. The QB928 strain is widely used in genetic studies due to the presence of various markers [aroI(aroK)906 purE1 dal(alrA)1 trpC2].
Several noncoding RNAs have been characterized in the B. subtilis genome in 2009, including Bsr RNAs. Microarray-based comparative genomic analyses have revealed that B. subtilis members show considerable genomic diversity.
FsrA, a small RNA found in B. subtilis, is an effector of the iron-sparing response and acts to down-regulate iron-containing proteins in times of poor iron bioavailability.
A promising fish probiotic, B. subtilis strain WS1A, possesses antimicrobial activity against Aeromonas veronii and suppressed motile Aeromonas septicemia in Labeo rohita. The de novo assembly resulted in an estimated chromosome size of 4,148,460 bp, with 4,288 open reading frames. B. subtilis strain WS1A genome contains many potential genes, such as those encoding proteins involved in the biosynthesis of riboflavin, vitamin B6, and amino acids (ilvD) and in carbon utilization (pta).
Conditional gene essentiality Studies using computational metabolic modeling have shown that gene essentiality in B. subtilis can vary depending on environmental conditions. Flux balance analysis simulations indicate that while some genes remain consistently essential for growth, others exhibit condition-dependent essentiality under nutrient or oxygen limitation.
Under nutrient-limited conditions, such as glucose or nitrogen restriction, metabolic activity shifts between biomass production and maintenance processes, altering growth outcomes. These findings suggest that metabolic robustness in B. subtilis involves tradeoffs between rapid growth and long-term survival under environmental stresses.
In response to nutrient limitation, B. subtilis can also initiate adaptive physiological responses, including metabolic diversification and endospore formation under prolonged stress.
Transformation Natural bacterial transformation involves the transfer of DNA from one bacterium to another through the surrounding medium. In B. subtilis, the length of transferred DNA is greater than 1,271 kb (more than 1 million bases). The transferred DNA is likely double-stranded DNA and is often more than a third of the total chromosome length of 4,215 kb. It appears that about 7–9% of the recipient cells take up an entire chromosome.
For a recipient bacterium to bind, take up exogenous DNA from another bacterium of the same species, and recombine it into its chromosome, it must enter a special physiological state called competence. Competence in B. subtilis is induced toward the end of logarithmic growth, especially under conditions of amino-acid limitation. Under these stressful conditions of semistarvation, cells typically have just one copy of their chromosome and likely have increased DNA damage. To test whether transformation is an adaptive function for B. subtilis to repair its DNA damage, experiments were conducted using UV light as the damaging agent. These experiments led to the conclusion that competence, with uptake of DNA, is specifically induced by DNA-damaging conditions, and that transformation functions as a process for recombinational repair of DNA damage.